Recent genetic analysis of parasite virulence confirms that there is an important link between increased parasite burden and disease caused by the AIDS pathogen, Toxoplasma gondii. The factors that control parasite burden are not understood, but it is clear that the balance between tachyzoite replication and switching to the persistence tissue cyst is critical to parasite numbers in the host. Gene transcription is an important mechanism in the control of Toxoplasma gondii development as evidenced by the large coordinate changes in transcription associated with parasite stage transitions that lead to the tissue cyst. We estimate that the number of genes under developmental control in the intermediate life cycle is ~20% of total gene expression detected in this parasite. Gene expression also varies between genetically diverse strains and these differences underlie phenotypic differences in parasite intracellular replication, tissue migration and invasion, virulence, and the establishment of long-term persistence. In this proposal, we will investigate the genetic basis for differences in the switching mechanism that accompanies formation of the bradyzoite stage found in the tissue cyst. In Aim 1, we will construct a comprehensive profile of tachyzoite- to-bradyzoite mRNA expression in early passage lines representing the three canonical lineages (Type I- GT-1, Type II-Me49B7, and Type III-CTG). Microarray data from a detailed kinetic series of all three strains induced to differentiate in vitro will be compared to profiles of brain cyst-derived bradyzoites in order to construct a high quality profile of tachyzoite-to-bradyzoite regulated mRNAs. These RNA profiles will also be compared with data from several bradyzoite mutants and from alternate methods of induction and host cell types in order to improve the overall resolution of these data. All hybridization results generated by this project will be downloaded onto ToxoDB in a format accessible to other investigators. In Aim 2, we will examine the genetic basis of developmental variation in a set of independent progeny obtained from a genetic cross between Type I-GT-1 and Type III-CTG. We will utilize microarray data to generate global mRNA profiles for genetic progeny under conditions that induce bradyzoite-specific gene expression. Differentially expressed mRNAs will be identified by standard methods, and then quantitative expression values and ratios from these analyses will be utilized as traits in QTL analysis to define chromosome regions and their interactions that control the expression of specific genes. These studies have the potential to identify genes that reduce developmental efficiency in Type I strains, while at the same time revealing groups of genes that positively regulate bradyzoite switching in more developmentally active strains. Recent genetic analysis of parasite virulence confirms that there is an important link between increased parasite burden and disease caused by the AIDS pathogen, Toxoplasma gondii. The factors that control parasite burden are not understood, but it is clear that the balance between tachyzoite replication and switching to the persistence tissue cyst is critical to parasite numbers in the host. In this proposal, we will investigate the genetic basis for differences in the switching mechanism that accompanies formation of the bradyzoite stage found in the tissue cyst, which is responsible for chronic toxoplasmosis. [unreadable] [unreadable] [unreadable]